Enzymatic production of biofuels (biodiesel) is generally conducted in multiphasic systems, and is a complex process. The reaction is a transesterification/esterification reaction, in which a fatty acid source (e.g. oil) and an alcohol or alcohol donor, are reacted in the presence of a lipase (or phospholipase) preparation, specifically immobilized lipase/phospholipase preparation, as disclosed, for example in applicant's WO11/107,977 and co-pending international patent application PCT/IL2011/000699.
Immobilization of enzymes has been described by a vast number of techniques basically aiming at reducing the cost contribution of enzymes in the overall enzymatic process; facilitating recovery of enzymes from the products; and enabling continuous operation of the process. Also the above WO11/107,977 and co-pending PCT/IL2011/000699 disclose techniques for immobilizing lipases/phospholipases. Generally, the immobilization techniques employ physical adsorption of enzymes to solid supports, such as silica and insoluble polymers; adsorption on ion-exchange resins, covalent binding of enzymes to a solid support material, such as epoxidated inorganic or polymeric supports, entrapment of enzymes in a growing polymer, confinement of enzymes in a membrane reactor or in semi-permeable gels or cross-linking enzyme crystals (CLECS's) or aggregates (CLEAS's). A main issue is to produce immobilized enzyme preparations which would be stable, whilst effective, so as to be used over a large number of reaction cycles, since the immobilized enzymes are expensive, and a cost-affecting parameter in all method of production using them.
Lipases and phospholipases exhibit low tolerance towards hydrophilic substrates, in particular short-chain alcohols and short-chain fatty acids (below C4). It has been observed in many research studies that short-chain alcohols and short-chain fatty acids, such as methanol and acetic acid, respectively, are responsible for detaching essential water molecules from the quaternary structure of those enzymes, leading to their denaturation and consequently loss of their catalytic activity. This drawback has prohibited the application of lipases for production of commercial quantities of fatty acids methyl esters (“biodiesel”) using oil triglycerides and methanol as substrates.
In this above described reaction of transesterification/esterification of a fatty acid source with a free alcohol, the formed glycerol and water by-products generally accumulate on the biocatalyst and/or its vicinity, blocking the substrates from free access to the active site of the immobilized enzyme. Such biocatalysts generally lose their catalytic performance after a few cycles when the same batch of biocatalyst is used. The special immobilized enzyme preparations, exhibiting good stability over many production cycles, persisting activity. Examples of such enzyme preparations are disclosed, inter alia, in applicant's WO/2008/084470, WO/2008/139455 and WO2009/069116.
Conditions under which the catalytic reaction is carried out, can adversely affect the stability and efficiency of immobilized enzyme preparations. It is important to have enzyme preparations which retain stability and activity under the reaction conditions.
Considering the factors which determine the reaction rates, the possibility to re-use the enzymes, etc., some of which are described above, the choice of the reactor is important. Applicant's WO11/107,977 and co-pending PCT/IL2011/000699 discloses a stirred tank reactor (STR), in order to obtain high yield and maintain the stability of the immobilized enzymes preparations.
Most biodiesel production studies with immobilized enzymes have reported the use of stirred tank reactors (STR), operated batchwise or in a continuous mode. Immobilized enzymes are mechanically stirred in a tank reactor containing a screen for retaining the enzyme for multiple use. Such a system has been found to be useful for achieving dispersion of the immobilized enzyme in the reactor; however due to high shear, resin-immobilized enzymes might be susceptible to attrition, leading to loss of the enzyme activity. Very few research studies have studied the use of immobilized enzymes also in a packed bed reactor (PBR). The glycerol byproduct, formed in the transesterification/esterification reaction was removed periodically from the PBR to prevent clogging the system. Such a system has been used at laboratory scale, but not at large or industrial scale where high pressure drop can be developed over the PBR which leads to inhibition of the continuous operation of the PBR.
In order to improve mass transfer for substrates and also avoid pressure drop in the system, co-solvents have been used by different work studies.
Other types of reactors including fluidized bed reactor (FBR), bubble column reactor (BCR) and expanded bed reactor (EBR) have not been evaluated nor suggested for the production of biodiesel with the aid of immobilized enzymes, and in any case such reactors are conventionally considered unsuitable for this purpose due problems associated therewith, including for example low conversions and loss of catalytic activity in the product stream (Sotoft et al., 2010, ibid.).